PROJECT SUMMARY Case reports of Zika virus (ZIKV) sexual transmission are mounting. Early epidemiologic studies demonstrate a higher incidence of ZIKV in women and also appear to demonstrate regional disparities in the clinical severity of Zika disease. These latter findings combined with laboratory reports indicate that prior immunity to dengue virus (DENV) or other related viruses and vaccines may modulate subsequent ZIKV infection. ZIKV persists in semen for up to 6 months and the vagina for up to 14 days. The most devastating outcomes of human ZIKV infection result from transmission of the virus from women to their fetuses. Thus, vaginal immunity emerges as a critical barrier to both stop the ZIKV transmission cycle, and to prevent a step towards eventual transmission to the fetus. Yet little is known about which mechanisms contribute to vaginal anti-ZIKV immunity and the roles that preexisting immunity to ZIKV or DENV play against subsequent vaginal ZIKV infection (vagZIKV). Recent studies from our lab and others using mouse models demonstrated that ZIKV replicates in the mouse vagina and that even a wild-type vaginal antiviral response is initially incompetent in controlling vagZIKV. Because DENV and ZIKV are highly cross-reactive and vaginal immunity is mediated in part by antibodies, it seems reasonable to speculate that antibodies would mediate cross-protection in the vagina. However, our recent data demonstrate that DENV/ZIKV-cross-reactive CD8 T cells dominate during subsequent systemic ZIKV infection, and that DENV-immune CD8 T cells are both necessary and sufficient to provide cross-protection against systemic ZIKV infection. Therefore, the goals of this project are (i) to evaluate the role of the CD8 T cell response in protection against vagZIKV. (Specific Aim 1) and (ii) To investigate how changes in prior ZIKV/DENV infection route/dose affect subsequent anti-ZIKV CD8 T cell responses, including primary effectors and various memory subsets (effector, central, and resident memory), and protection against vagZIKV (Specific Aim 2).